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Am J Hum Genet. 2018 Oct 4;103(4):498-508. doi: 10.1016/j.ajhg.2018.07.016. Epub 2018 Sep 12.

A Multiplex Homology-Directed DNA Repair Assay Reveals the Impact of More Than 1,000 BRCA1 Missense Substitution Variants on Protein Function.

Author information

1
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA; Brotman Baty Institute for Precision Medicine, Seattle, WA 98195, USA.
2
Department of Biomedical Informatics, Ohio State University, Columbus, OH 43210, USA.
3
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA.
4
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA; Department of Medicine, University of Washington, Seattle, WA 98195, USA; Howard Hughes Medical Institute, Seattle, WA 98195, USA.
5
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA; Brotman Baty Institute for Precision Medicine, Seattle, WA 98195, USA; Howard Hughes Medical Institute, Seattle, WA 98195, USA. Electronic address: shendure@u.washington.edu.
6
Department of Biomedical Informatics, Ohio State University, Columbus, OH 43210, USA; Comprehensive Cancer Center, Ohio State University, Columbus, OH 43210, USA. Electronic address: jeffrey.parvin@osumc.edu.

Abstract

Loss-of-function pathogenic variants in BRCA1 confer a predisposition to breast and ovarian cancer. Genetic testing for sequence changes in BRCA1 frequently reveals a missense variant for which the impact on cancer risk and on the molecular function of BRCA1 is unknown. Functional BRCA1 is required for the homology-directed repair (HDR) of double-strand DNA breaks, a critical activity for maintaining genome integrity and tumor suppression. Here, we describe a multiplex HDR reporter assay for concurrently measuring the effects of hundreds of variants of BRCA1 for their role in DNA repair. Using this assay, we characterized the effects of 1,056 amino acid substitutions in the first 192 residues of BRCA1. Benchmarking these results against variants with known effects on DNA repair function or on cancer predisposition, we demonstrate accurate discrimination of loss-of-function versus benign missense variants. We anticipate that this assay can be used to functionally characterize BRCA1 missense variants at scale, even before the variants are observed in results from genetic testing.

KEYWORDS:

BRCA1; DNA repair; VUS; missense variants; variants of uncertain significance

PMID:
30219179
PMCID:
PMC6174279
DOI:
10.1016/j.ajhg.2018.07.016
[Indexed for MEDLINE]
Free PMC Article

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